In vitro, the effects of BM-1 (extracts from the mackerel) on platelet aggregation induced in platelet rich plasma (PRP) by ADP, collagen, epinephrine and A. A. (arachidonic acid) was inhibited by BM-1. But platelet aggregation by PMA (phorbol 12, 13-myristate acetate) was not inhibited by BM-1.
In the platelet suspension, when BM-1 was removed from the medium, inhibition of platelet aggregation by BM-1 was not observed.
Cytoplasmic free Ca²⁺ was measured in human platelets using the intracellularlytrapped fluorescent indicator Fura-2.
Thrombin-induced increase in cytoplasmic free Ca²⁺ was inhibited by BM-1. When BM-1 was removed from the medium, inhibition of the rise in cytoplasmic free Ca²⁺ was not found.
The present findings would suggest that there were unknown factors in BM-1 which inhibited the aggregation of platelet and the rise in cytoplasmic free Ca²⁺ of platelets.

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Plasma levels of vitamin K, which is composed of phylloquinone and menaquinone and protein C antigen were measured in thirty patients on long-term warfarin therapy after cardiac surgery. Thirty patients were devided in two groups by means of the type of cardiac surgery. Twenty patients of group A were the cases of cardiac valve replacement and ten patients of group B were the cases of coronary artery bypass grafting. Group A was divided further in two grops, group A-(1) and group A-(2) according to the amount of warfarin doses. Vitamin K was measured by high-performance liquid chromatography and protein C antigen was measured by ELISA method.
Thrombotest values of groups A and B ranged on an average of 20.2±3.9% and 20.7±5.3%. Plasma levels of vitamin K and protein C antigen in both groups were on an average of 1.36±0.44ng/ml versus 2.64±1.56ng/ml (p<0.05) and 93.4±20.22% versus 107.2±13.94% respectively. There was no difference between group A-(1) and A-(2).
There was no case in which vitamin K and protein C decreased below the normal levels. These findings suggest that long-term warfarin therapy has no influence on the synthesis of vitamin K and protein C in patients after cardiac surgery.

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The thrombolytic efficacy of plasminogen activators (PAs) were compared using juglar vein thrombus model of rabbits (Collen et al. J. Clin. Invest. 71, 368-376, 1983). PAs were one-chain, two-chain and the mixure of both chains of recombinant tissue PAs (one-, two- and m-rTPAs) and high molecular weight of urokinase (UK). The thrombus was formed using ¹²⁵I-labeled human fibrinogen. The thrombolytic ratio was calculated by the radioactivities of thrombus, circulating blood, washing water surround the thrombus and the catheter used for washing out the free radioactivity in the external juglar vein ligated at the time of thrombus formation. The levels of PAs were measured by Labeled Avidin-Biotin technique using anti-TPA polyclonal antibody and bioninated anti-TPA antibody. The levels of blood elements concerning with thrombolysis were also measured at two hours intervals.
1. The thrombi were dissolved dose-dependently from 0.055 to 0.2mg/kg body weight of m-rTPA and the mamimum thrombolytic ratio was calculated 72±9% (n=5). The maximum ratio of one- and two-rTPAs was the same value of m-rTPA. The concentration of these rTPAs was considered around 0.4mg/kg at the maximum ratio. On the other hand, the maximum thrombolytic ratio using by UK was remained around 43±11% (n=6) and needed concentration of UK to obtained the maximum ratio was around 2mg/kg.
2. The levels of fibrinogen, plasminogen and α₂-plasmin inhibitor in circulating blood were not decreased using rTPAs, but decreased using UK dose-dependently. No significant difference of these parameters was recognized among the differences of rTPAs employed.
3. 10% of total volume of rTPAs were administered as a bolus injection and the residual 90% were administered as continuous infusion over 4 hours and the levels were measured. Although it was found a tendency that the level of one-rTPA in plasma was higher than that of two-rTPA, the difference was not significant statistically.
It was concluded that rTPAs appears to be superior to UK, because of its higher (over 10 fold) specific thrombolytic efficacy than that of UK. In this thrombus model experiment using rabbits, no difference was found in thrombolitic efficacy and effect on coagulation parameters in plasma among three different rTPAs.

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Plasma level of cross-linked fibrin degradation products (XDP) was measured by latex agglutination using a new monoclonal antibody against Decate (Iatron Co., Tokyo, Japan). Plasma XDP (expressed as DD/E) was undetectable (<1μg/ml) in 80 healthy subjects (mean age 43.7±19.1 years), but showed high levels in 14 of 15 patients with DIC. The levels of DD/E and total FDP measured with antibody against human fibrinogen changed in parallel with their clinical course during treatment with heparin for DIC. Furthermore, there was significant correlation between levels of DD/E and total FDP in 46 samples from patients with DIC (p<0.01). But the level of total FDP was very high in comparison with that of DD/E in a DIC patient, and in 3 patients with acute myocardial infarction during infusion of recombinant t-PA. Those results suggested that this method for XDP was sensitive and useful to assessment of fibrinolysis in circulating blood.

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The relationship between tissue plasminogen activator (tPA), its fast acting inhibitor (PAI-1) and euglobulin clot lysis assay (ELT) were investigated with healthy volunteers' plasma.
New clot lysis assay method was utilized with a slight modification for ELT. Briefly, clot was made in a microtiter plate and turbidity of the clot was measured with 340nm by a microtiter plate reader periodically. The point of half lysis was determined as euglobulin lysis time 1/2 (Fig. 1). Duplicated samples were analyzed and the correlation coefficient (R) was 0.992 (p<0.001). TPA was assayed by EIA.
Total PAI (PAIt) and tPA-PAI complex (PAIc) were measured separately by the methods previously reported from this laboratory. Both tPA and PAI showed the significant correlation with ELT. TPA had a significantly positive, not negative, correlation with ELT (R=0.387, p<0.001). Higher correlation coefficients (R=0.580, p<0.001 and R=0.599, p<0.001) were obtained with PAIt and PAIf than with tPA and PAIc (R=0.427, p<0.001). The positive correlation was also obtained between tPA and PAI. PAIt and PAIc showed higher correlation coefficients (R=0.733, p<0.001 and R=0.827, p<0.001) with tPA than with PAIf (R=0.648, p<0.001). These data suggest that PAI can be released corresponding to the release of tPA and that the activity of tPA can be neutralized by PAI. That can be the reason why PAIt and PAIf were the more important factor to influence the ELT than tPA. We here show that PAI is highly important factor for ELT, especially, the amounts of free PAI being the key factor to determine the ELT, which can represent the potential activity of the fibrinolytic system.

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Plasminogen activator inhibitor (PAI) activity was measured by a newly developed assay (PAI-test, Behringwerke AG) in 17 healthy subjects, 52 patients with thrombotic disease, seven patients with sepsis and three pregnent women. This assay method uses an oxidant which destroys α₂-plasmin inhibitor activity, and acidification and dilution steps of plasma samples could be eliminated. The mean PAI activity was 0.67±SD0.60 u-PA inhibiting units/ml (U/ml) in healthy subjects; men had somewhat higher mean PAI levels than women (0.81 vs 0.54U/ml). Some patients with thrombotic diseases had elevated PAT values (1.04±1.09U/ml), and their PAI values correlated with tissuetype plasminogen activator (t-PA) antigen but not with plasmin-α₂-plasmin inhibitor complex. Increased PAI levels were found in patients with sepsis (3.66±3.22U/ml) and in pregnant women (7.80±4.45U/ml), Further studies of larger series of patients would provide more precise information about the clinical implication of high PAI states.

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The presence of an inhibitor against factor VIII or factor IX must still be seen as the most important complication in the treatment of hemophilia patients. Brackmann et al. reported successfully induced immune tolerance for hemophilia patients with inhibitor. We tried induction immune tolerance for hemophilia patient with inhibitor.
A hemophilia A patient (10 years old boy) with high level of inhibitor has treated with regular transfusion of low dose factor VIII. Over 30 months factor VIII (about 23u/kg) has transfused 3 times a week. About 6 month later of the beginning treatment the level of inhibitor level has fallen to low level (1.1 Bethesda units) and factor VIII in vivo recovery has improved. The level of inhibitor shows low level over 24 months and bleeding episodes has dramaticaly reduced. The patient is successfully induced immune tolerance and his school life has improved.

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We succeeded in an anticoagulant therapy of a postcaval giant thrombus in a 14 year old male with a heterozygous AT III deficiency. The diagnosis of AT III deficiency was confirmed by laboratory findings of him and his family members on the neneth hospital week (Table 1, 2, Fig. 1). He was initially treated with gabexate mesilate, and then with AT III concentrate and warfarin. These treatments brought remarkable improvement of his clinical status and labolatory findings (Fig. 2). Four months after initiation of the treatment with AT III concentrate and warfarin, the thrombus of postcava disappeared. These results suggested that therapy with AT III concentrate and warfarin was effective not only in acute phase but in subacute phase of thrombosis in AT III deficiency.
Furthermore we examined FDP-E, FDP-D dimer, TAT complex and plasmin-α₂PI complex in nine relatives with heterozygous AT III deficiency who had not experienced thrombotic problems (Table 3). Abnormal values were observed in eight of nine on TAT complex, seven of nine on Plm-α₂PI complex, four of nine on FDP-E, and two of nine on FDP-D dimer. These data revealed latent hypercoagulable states in some of these patients. And consequently, analysis of these markers might be useful to detect the risk of thrombosis in AT III deficiency.

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